The objective of the proposed research is the purification, physical and chemical characterization of the 450,000 dalton glycoprotein (TSN) obtained from the supernatant produced by centrifuging out human blood platelets after thrombin treatment of the washed cells. This molecule appears to consist of three main chains of 150,000 daltons which are disulfide linked, as well as a variable complement of light chains which are removed by high ionic strength treatment. The characterization of proteolytic fragments of TSN, particularly a core particle known to be produced by trypsin is expected to assist in the structural studies, and possibly in determining the relationship between TSN and the clotting scheme. Investigation of the effects of proteolysis, factors 9a, 10a, 11a, and 12a may indicate that TSN is a highly specific substrate. TSN binds heparin relatively weakly. The strength of this interaction will be measured against affinity for other polysaccharides such as dermatan sulfate or chondroitin sulfate. Attempts will be made to relate TSN to known coagulation functions and to confirm its relationship to a specific cytoplasmic granule fraction in the platelet. Preliminary experiments indicate that highly purified TSN will allow the production of a potent and specific antiserum which will be useful in antibody labelling studies concerned with the origin and destiny of TSN and may eventually be valuable clinically. Mapping the TSN molecule by electron microscopy will permit the determination of molecular shape and size; complexes of TSN protein markers specific for particular sites (such as loci of light chain attachment, glycosidic chains, or the heparin binding site, using IgG, lectins, and labelled heparin) will be visualized to determine specific sites with respect to the general topography of the molecule.